PROJECT SUMMARY/ ABSTRACT Although cognitive decline (CD) and impairment are widespread and exact an enormous healthcare burden, therapeutic options are limited partly due to our incomplete understanding of the neurobiological bases of CD. Across conditions, including Alzheimer's, and Parkinson's Disease, worsening CD is commonly associated with high activity of the catechol-O-methyltransferase (COMT) enzyme. Such high activity can occur as a consequence of genetic variation in the COMT gene. The most studied COMT genetic polymorphism, rs4680, is an A?G change that yields a 4-fold difference in COMT enzyme activity. COMT breaks down catecholamines, and is the primary regulator of dopamine clearance in cortical brain regions; it also detoxifies carcinogenic catechol-estrogens. There are two COMT isoforms, which encode soluble (S-COMT) and membrane bound (MB-COMT) forms of the enzyme, respectively. Interventions that inhibit COMT can improve cognitive function, presumably by elevating cortical dopamine. However, while inhibiting MB-COMT, which dominates in the brain, shows cognitive benefit, no COMT inhibitors to date are isoform selective, and inhibition of S-COMT increases circulating carcinogenic catechol-estrogens, as well as peripheral catecholamines, increasing hormone-sensitive cancer risk, and pain sensitivity, respectively. While the evidence is strong that COMT genetic variation alters enzyme function, growing evidence also points to the influence of epigenetic COMT regulation, e.g., DNA methylation, in moderating COMT gene expression and bulk enzyme activity. Thus, the ability to selectively downregulate MB-COMT expression holds tremendous promise as a therapeutic intervention for CD across a broad array of conditions. Nevertheless, major mechanistic gaps persist: 1) There currently is no efficient way to distinguish and accurately measure MB- and S-COMT transcripts, and COMT protein isoforms haven't been measured in tandem with COMT mRNAs; 2) DNA methylation remains to be completely interrogated throughout regions thought to regulate MB- and S-COMT mRNA expression; and 3) the metabolites driving observed epigenetic modification of COMT, and their isoform specificity, are unknown. We therefore propose an in vitro study as a necessary first step in understanding the effects of metabolites and other epigenetic regulators on isoform-specific COMT expression in human cell lines. We expect to identify compounds regulating COMT, and their underlying epigenetic mechanisms, which will identify both biomarkers of CD risk and targets for new interventions for CD. Such future interventions may be particularly useful in combating executive function deficits, which is an aspect of CD common to a broad array of conditions, including normal aging, multiple neurodegenerative conditions, schizophrenia, ADHD, PTSD, depression, substance use disorders, and cancer chemotherapy treatment. As epigenetic marks can be reversed, a better understanding of isoform-specific regulation of COMT will be transformative in identifying new strategies for treating and delaying CD in these populations, substantially reducing their clinical and public health burden.